Antimicrobial substrates and uses thereof

ABSTRACT

A new substrate makes it possible to modify surface properties relating to antimicrobial properties. Said substrate has an electron donating surface, characterized in having metal particles on said surface, said metal particles comprising palladium and at least one metal chosen from gold, ruthenium, rhodium, osmium, iridium, and platinum, wherein the amount of said metal particles is from about 0.001 to about 8 μg/cm 2 . The substrate is suggested for different uses, such as for modifying the hydrophobicity, protein adsorption, adhesion of bacteria, as well as preventing bacterial transmission and in particular preventing nosocomial infections.

RELATED APPLICATION

This application claims priority to U.S. Provisional Application Ser.No. 60/790,307 filed Apr. 7, 2006, which is hereby incorporated byreference in its entirety.

FIELD OF THE INVENTION

The present invention relates to a new antimicrobial substrate with nanoparticles, which makes it possible to modify surface properties relatingto antimicrobial properties in a repeatable and controlled manner.Examples of surface properties, which can be modified, include but arenot limited to hydrophobicity, protein adsorption, and adhesion ofbacteria. Examples of uses of the present substrate include but are notlimited to delaying colonisation of bacteria, preventing transmission ofbacteria and in particular nosocomial infections. The present inventionfurther relates to objects comprising said new substrate. The presentinvention further relates to the use of said substrate. Finally thepresent invention relates to a method for the manufacture of such asubstrate.

BACKGROUND

It has always been desirable to modify surface characteristics toachieve useful properties. In particular it is desired to be able tomodify surface properties that are important in connection withantimicrobial objects.

SHORT SUMMARY OF THE PRESENT INVENTION

A problem in the state of the art regarding surfaces is how to provide asurface which for example is antimicrobial, wherein it in a repeatableway is possible to modify the hydrophobicity, protein adsorption, andadhesion of bacteria.

The present inventors have discovered that the above-mentioned problemin the state of the art is solved by a substrate having an electrondonating surface, characterized in that there are metal particles on thesurface. The metal particles include palladium and at least one metalchosen from gold, ruthenium, rhodium, osmium, iridium, and platinum andwherein the amount of said metal particles is from about 0.001 to about8 μg/cm². Further embodiments of the present invention are defined inthe appended dependent claims.

DESCRIPTION Definitions

Before the invention is disclosed and described in detail, it is to beunderstood that this invention is not limited to particularconfigurations, process steps and materials disclosed herein as suchconfigurations, process steps and materials may vary somewhat. It isalso to be understood that the terminology employed herein is used forthe purpose of describing particular embodiments only and is notintended to be limiting since the scope of the present invention islimited only by the appended claims and equivalents thereof.

It must be noted that, as used in this specification and the appendedclaims, the singular forms “a”, “an” and “the” include plural referentsunless the context clearly dictates otherwise.

The following terms are used throughout the description and the claims.

“Adhesion of bacteria” as used herein describes the phenomenon wherebacteria adhere to a surface.

“Antimicrobial” as used herein encompasses the property of suppressingand/or eliminating microbial growth.

“Colonisation” as used herein encompasses the establishment of coloniesof for instance bacteria.

“Contact angle”. For a given droplet on a solid surface the contactangle is a measurement of the angle formed between the surface of asolid and the line tangent to the droplet radius from the point ofcontact with the solid.

“Electron donating material” as used herein encompasses a material,which in connection with another more noble material has the ability totransfer electrons to the more noble material. An example is a lessnoble metal together with a more noble metal.

“Electron donating surface” as used herein encompasses a surface layercomprising an electron donating material.

“Hydrophobicity” of a surface as used herein describes the interactionsbetween the surface and water. Hydrophobic surfaces have little or notendency to adsorb water and water tends to “bead” on their surfaces.The term hydrophobicity of a surface is also closely linked with itssurface energy. Whereas surface energy describes interactions of thesurface with all molecules, the hydrophobicity describes theinteractions of the surface with water.

“Hysteresis of contact angle” as used herein is the difference betweenthe advancing and receding contact angle values. The advancing contactangle of a drop of water on a surface is the contact angle when theboundary between water and air is moving over and wetting the surface,while the receding angle is the contact angle when boundary betweenwater and air is withdrawn over a pre-wetted surface.

“Modify” either means reducing or enhancing a property.

“Noble” is used herein in a relative sense. It is used to relatematerials including metals to each other depending on how they interactwith each other. When two metals are submerged in an electrolyte, whileelectrically connected, the term “less noble” metal is used to denotethe metal which experiences galvanic corrosion. The term “more noble” isused to denote the other metal. Electrons will be transferred from the“less noble” metal to the more noble metal.

“Nosocomial infection” as used herein describes an infectious diseasespreading for instance in a hospital environment.

“Protein adsorption” as used herein encompasses the phenomenon whereproteins adhere to a surface due to overall attractive forces betweenthe proteins and the surface.

“Substrate” as used herein is the base, which includes the material thatis treated according to the present invention.

DETAILED DESCRIPTION OF THE PRESENT INVENTION

According to the present invention a substrate is treated to give itdesired properties. The substrate can be made of a wide range ofmaterials. In one embodiment the substrate is made of a material, whichhas an electron-donating surface. In the case of an electron-donatingsurface the metal particles can be applied directly on to theelectron-donating surface. In the case where the surface it not electrondonating, a layer of an electron donating material has to be applied tocreate an electron donating surface.

The present disclosure describes a substrate having an electron donatingsurface. The substrate includes metal particles on said surface. Themetal particles include palladium and at least one metal chosen fromgold, ruthenium, rhodium, osmium, iridium, and platinum. The amount ofmetal particles is from about 0.001 to about 8 μg/cm² on the surface. Apreferred amount of the metal particles is from about 0.01 to about 4μg/cm². A particularly preferred amount of the metal particles is fromabout 0.01 to about 1 μg/cm².

Either the substrate itself is electron donating or there is applied alayer of an electron donating material on the substrate. In the casewhere the electron donating material is applied on the substrate it isapplied in an amount of from about 0.05 to about 12 μg/cm².

An electron donating material does not necessarily have anelectron-donating surface. An example is aluminium, which in air gets anoxide layer, which is not an electron-donating surface.

The electron donating material is any material with the ability to forman electron-donating surface, such as a conducting polymer or a metal.In the case of a metal it must be less noble than any of the metals inthe group consisting of palladium, gold, ruthenium, rhodium, osmium,iridium, and platinum.

A preferred metal for use as an electron-donating surface is a metalchosen from silver, copper and zinc.

In one embodiment of the present invention the substrate is a polymericsubstrate.

In one embodiment the substrate is chosen from latex, polymerscomprising vinyl groups, silicone, polyvinylchloride, polypropylene,polyurethane, polyester, copolymerisates of ethylene vinyl acetate,polystyrene, polycarbonate, polyethylene, polyacrylate,polymethacrylate, acrylonitrile butadiene styrene, polyamide, andpolyimide, or mixtures thereof.

In another embodiment of the present invention the substrate is chosenfrom a natural polymer, a degradable polymer, an edible polymer, abiodegradable polymer, an environmental friendly polymer, and a medicalgrade polymer.

In another embodiment of the present invention the substrate is a metal.

A preferred metal for the substrate is chosen from stainless steel,medical grade steel, titanium, medical grade titanium, cobalt, chromiumand aluminium or mixtures thereof.

In another embodiment of the present invention the substrate is chosenfrom glass, minerals, zeolites, stone and ceramics.

In another embodiment of the present invention the substrate is chosenfrom paper, wood, woven fibres, fibres, cellulose fibres, leather,carbon, carbon fibres, graphite, polytetrafluoroethylene, andpolyparaphenyleneterephthalamide.

In another embodiment of the present invention the substrate has theshape of a particle. In this embodiment particles are coated accordingto the present invention.

Such particles can have a spherical shape or an irregular shape.

In one embodiment of the present invention there is provided an objectcomprising a substrate according to the present invention. Examples ofan object comprising a substrate according to the present invention aremedical devices, medical instruments, disposable articles, medicaldisposable articles.

The particles must always include palladium. In addition to palladiumthere is at least one other metal. A ratio of palladium to other metalsin the metal particles of from about 0.01:99.99 to about 99.99:0.01 canbe used in the present invention. A ratio from about 0.5:99.5 to about99.8:0.2 is preferred. Particularly preferred ratios are from about 2:98to about 95:5. Very particularly preferred ratios are 5:95 to 95:5. Inanother embodiment the ratios are from about 10:90 to about 90:10.

In one embodiment of the present invention the metal particles, inaddition to palladium, include gold.

The present inventors have discovered that advantageous properties areachieved when the metal particles have an average size of from about 10to about 10000 Å.

In one embodiment the average sizes for said metal particles are fromabout 100 to about 600 Å.

In another aspect of the present invention there is provided an objectincluding any of the substrates described herein.

There is also provided a medical device comprising any of the substratesdescribed herein.

A disposable article comprising any of the substrates described hereinis also provided.

The present invention also provides a dental article, as well as dentalequipment, dental implants, and dental devices, comprising any of thesubstrates described herein.

The applied amount of the metal particles is expressed as a surfaceconcentration in μg/cm² and it must be realised that the metal particlesdo not form a covering layer, but instead are uniformly distributedparticles or clusters on said electron donating surface. Thus this is ameasure of the weight of the particles on an area of the substrate.

An applied layer of an electron donating material is preferably appliedso that it is uniform, essentially without agglomerates or clusters onthe surface. If the electron donating surface layer is homogenous anduniform the applied amount in μg/cm² may be converted to a thickness inA. An applied amount of 0.05-4 μg/cm² corresponds to about 4.8-380 Å,0.5-8 μg/cm² corresponds to about 48-760 Å, and 0.8-12 μg/cm²corresponds to about 76-1140 Å.

In one embodiment of the present invention the electron-donating surfaceis a layer of commercially available essentially pure silver, which doesnot exclude the possibility of small amounts of impurities.

If the substrate does not have an electron donating surface and thus adeposition of an electron donating surface layer is necessary, thedeposition is performed using a method chosen from chemical vapourdeposition, sputtering, and deposition of metal from a solutioncomprising a metal salt. A uniform layer essentially without clusters oragglomerates is the result of the deposition. Preferably the depositionis carried out so that the first layer has good adhesion to thesubstrate.

Now there is described one embodiment of the present invention forpreparation of the coated substrate. For substrates which do not have anelectron donating surface the method includes some or all of thefollowing steps:

1. pre-treatment2. rinsing3. activation4. deposition of an electron donating surface5. rinsing6. deposition of metal particles7. rinsing8. drying

For objects with an electron-donating surface the method comprises thesteps

1. rinsing2. deposition of metal particles3. rinsing4. drying

In the following, one embodiment of steps 1 to 8 for substrates which donot have an electron-donating surface are described more in detail.

The pre-treatment can be made in an aqueous solution of a stannous saltcontaining from about 0.0005 to about 30 g/l of stannous ions. The pH isfrom about 1 to about 4 and adjusted by hydrochloric and/or sulphuricacid. The treatment time is from about 2 to about 60 minutes at roomtemperature. After the pre-treatment the surface is rinsed indemineralised water, but not dried.

The activated and rinsed substrate is transferred to the depositionsolution. The deposition solution has a pH of not less than about 8. Itincludes a metal salt chosen from a silver salt, a zinc salt, and acopper salt. In one embodiment of the present invention the salt issilver nitrate (AgNO₃). The metal salt is used in an effective amount ofno more than about 0.10 grams per litre, preferably about 0.015 gramsper litre. If the metal content is above about 0.10 grams per litre, theelemental metal may form nonuniformly, in the solution or on thecontainer walls. If the metal content is below an effective amount,there is insufficient metal to form a film in the desired time.

A second component of the deposition solution is a reduction agent thatreduces the metal-containing salt to elemental metal. The reductionagent must be present in an amount sufficient to accomplish the chemicalreduction.

Acceptable reduction agents include formaldehyde, hydrazine sulphate,hydrazine hydroxide, and hypo phosphoric acid. In one embodiment of thepresent invention it is present in an amount of about 0.001 millilitresper litre of solution. Too large a concentration of the reduction agentcauses deposition of metal throughout the solution and on the containerwalls, while too small a concentration may result in an insufficientformation of metal on the substrate. A person skilled in the art can inthe light of this description by routine experimentation determine thedesired amount of reduction agent.

Another component of the deposition solution is a deposition controlagent that is present in an amount sufficient to slow the depositionreaction to prevent the reduced metal from precipitating directly fromsolution as a fine metallic powder, or precipitating onto the walls ofthe container. Operable deposition control agents include invertedsugar, also known as invertose, succinic acid, sodium citrate, sodiumacetate, sodium hydroxide, potassium hydroxide, sodium tartrate,potassium tartrate, and ammonia. The deposition control agent ispreferably present in an amount of about 0.05 grams per litre ofsolution. If too little is present, there may occur precipitation ofmetal clusters instead of a uniform metallic surface. If too much ispresent, the metal-containing salt may become too stable for the desiredprecipitation onto the substrate of interest.

The concentrations of the reduction agent and the deposition controlagent are adjusted as necessary to achieve the desired results,depending upon the substrate material, the thickness of the filmdesired, the conditions of deposition, and the concentration of metal inthe solution. For example, for thin films the metal salt concentrationwill be relatively low, as will the concentrations of the reductionagent and the deposition control agent. A person skilled in the art canin the light of this description by routine experimentation determinethe desired amount of deposition control agent.

In preparing the deposition solution, each of the components of thesolution are preferably individually dissolved in demineralised water.The various pre-solutions are then mixed, and diluted where necessary,in the correct amounts to achieve the concentrations mentioned above.

The combination of a metal salt and reduction agent permits the metal tobe reduced from the salt in a suitable state to be deposited upon thesurface of the substrate. This method is particularly beneficial toachieve good adhesion of the completed metal film to the substratesurface. Good adhesion is important in nearly all uses.

The substrate surface is exposed to the deposition solution by anyappropriate procedure. Dipping into the solution is normally preferred,but the solution may be applied by any convenient technique such asspraying or brushing. The metal film deposits uniformly from thesolution at a rate that may be controlled by the concentration of themetal salt. If a thin film is required, the temperature of deposition ismaintained sufficiently low so that deposition is controllably slow.

Other methods of applying a metal layer that acts as anelectron-donating surface can also be applied in the present invention.Other ways of achieving an electron-donating surface include chemicalvapour deposition and sputtering.

After the above-described metal deposition the substrate has anelectron-donating surface consisting of a metal. This metal depositionis only necessary if the substrate does not have an electron-donatingsurface from the start. If the substrate already possesses anelectron-donating surface, metal particles can be deposited on thesurface without the extra addition of a metal layer. In the latter casethe substrate is cleaned thoroughly before application of the particles.

The next step in the manufacturing method is deposition of metalparticles.

In one embodiment colloidal suspensions of metals are used to obtainparticles comprising palladium and at least another metal on thesurface. The metal particles are deposited from a suspension of thedesired particles. The composition of the metal particles in thesuspension is adjusted according to the preferred value. The substratewith the electron-donating surface is dipped in the suspension of metalparticles for a period of time from about a few seconds to about a fewminutes or longer.

The suspension of metal particles can be manufactured in several ways.In one embodiment the suspension of metal particles is made from anaqueous solution of a metal salt which is reduced under conditions suchthat metal particles of a desired size are formed. Mixing a suitableamount of metal salt, reducing agent and stabilising agent achievesthis. The same reducing agents and stabilising agents as described abovecan be used when making the particle suspension. A person skilled in theart can in the light of this description by routine experimentationdetermine the desired amount of reducing agent and stabilising agent toget the desired particle size. In an alternative embodiment acommercially available colloidal suspension of metal particles is used.Metal particles of the desired composition are used to make thesuspension.

In one embodiment the suspension of metal particles is made by dilutingwith demineralised water a commercially available concentrated colloidalsolution of metal particles comprising palladium and at least one metalchosen from gold, ruthenium, rhodium, osmium, iridium, and platinum. Thesubstrate is treated with the suspension for a period of time from abouta few seconds to about a few minutes or longer. After the treatment thesubstrate is rinsed in a solvent or water such as demineralised waterand left to dry in room temperature.

In one particular non-limiting embodiment the commercially availablemetal particles consist of 75% palladium and 25% gold.

Thus according to the present invention, a substrate with a particulardesired surface can be obtained. For example, one can prepare asubstrate having a silver electron donating surface with particlesconsisting of 75% palladium and 25% gold, or a copper electron donatingsurface with particles consisting of 85% palladium and 15% ruthenium.

One of the advantages offered by the flexible yet controlled andrepeatable method for producing such substrates is that a wide varietyof substrates can be produced. As described further herein, certainsubstrates have improved properties over existing substrates. Forexample a particular substrate according to the present invention canproduce surprising and advantageous modifications of the hydrophobicityof a substrate to which is it applied. Other properties that can bemodified in this way by substrates according to claim 1 include proteinadsorption and adhesion of bacteria.

That is, it is possible to adjust the particle size, the composition ofthe particles and the amount of particles to modify the surfaceproperties of objects to which the substrate is applied.

The present inventors have discovered that it is possible to achievethis by using a substrate according to claim 1. In particular it ispossible to adjust the particle size, the composition of particles, andthe amount of particles to modify the surface properties.

Substrates according to the present invention can be used for manypurposes. They are suitable for use in any application where it isdesired to modify hydrophobicity, protein adsorption, and adhesion ofbacteria of a substrate.

Properties of the substrate can be both reduced or increased. Thusobjects are provided which display at least one area which enhances afeature, and at least one area which reduces a feature. An example is anobject with an area that reduces protein adsorption and an area thatenhances protein adsorption.

A substrate according to the present invention also comprises asubstrate having an electron donating surface, with metal particles onsaid surface, said metal particles comprise palladium wherein the amountof said metal particles is from about 0.001 to about 8 μg/cm².

The present invention provides use of a substrate according to thepresent invention for modifying the protein adsorption to an objectcomprising said substrate. An example of use is to adjust the proteinadsorption onto a medical device to a desired level.

The present invention provides use of a substrate according to thepresent invention for modifying the bacterial adhesion to an objectcomprising said substrate.

The present invention provides use of a substrate according to thepresent invention for preventing bacterial growth.

The present invention provides use of a substrate according to thepresent invention for preventing colonisation of bacteria.

An example of this where it is important to modify the adhesion ofbacteria, preventing bacterial growth and preventing colonisation ofbacteria is a catheter to be inserted in a body, where the adhesion ofbacteria, bacterial growth and colonisation of bacteria should be as lowas possible.

The present invention provides use of a substrate according to thepresent invention for preventing transmission of bacteria. Transmissionof bacterial infections is prevented by the prevention of thetransmission of bacteria. Examples of objects used in this context arehandles, buttons, switches, hospital equipment, surgical instruments,medical instruments, kitchen equipment, and all other objects, which areable to transmit bacteria.

The present invention provides use of a substrate according to thepresent invention for preventing transmission of a nosocomial infection.An object comprising a substrate according to the present invention canbe used in any context where it is desired to prevent transmission of abacterial infection. Preventing transmission of bacteria and thusbacterial infections will in particular prevent nosocomial infections.

Other features of the invention and their associated advantages will beevident to a person skilled in the art upon reading the description andthe examples.

It is to be understood that this invention is not limited to theparticular embodiments shown here. The following examples are providedfor illustrative purposes and are not intended to limit the scope of theinvention since the scope of the present invention is limited only bythe appended claims and equivalents thereof.

EXAMPLES Example 1 Hydrophobicity of the Surface as a Function of theAmount of Metal Particles

A uniform layer of silver was deposited on a glass substrate accordingto the following method. The substrate was immersed in a cleaningsolution of chromic acid for 5 minutes at 58° C., followed by rinsing indemineralised water. The surface of the substrate was activated byimmersion in a solution of aqueous stannous chloride and then rinsed indemineralised water. The surface of the substrate was then plated with auniform layer of silver by immersion in 3 deposition solutionscomprising silver ions. This yielded a silver surface with an appliedamount of 1.2 μg/cm² corresponding to a thickness of about 115 Å.

Particles consisting of 23% palladium and 77% gold were subsequentlydeposited on the first silver surface by immersion in a dilutesuspension comprising metal particles of gold/palladium. The suspensionof metal particles was made by reducing a gold salt and a palladium saltwith a reducing agent and stabilising the suspension with a stabilisingagent. The substrate was subsequently rinsed in demineralised water anddried.

Substrates with different amounts of deposited particles were made usingthe method outlined above. Amounts of particles were 0, 0.02, 0.11,0.15, and 0.19 μg/cm² respectively. For the sample with 0 μg/cm² noparticles were deposited on the surface and hence it consists of asilver surface.

The static contact angle of a drop of water in equilibrium on thedifferent substrates was measured. The advancing and receding contactangles were measured using the Wilhelmy technique.

The difference between the advancing and receding contact angle valuesis called the contact angle hysteresis and was calculated for themeasurements. The result of the experiment is depicted in Table 1.

TABLE 1 Amount of Static contact Contact angle particles anglehysteresis (μg/cm²) (degrees) (degrees) 0 52 70 0.02 50 77 0.11 56 750.15 62 80 0.19 62 84

The surface hydrophobicity of the substrate is thus modified while thesurface displays several other useful properties, such as antimicrobialproperties, inherent of the substrates according to this example.

Example 2 Protein Adsorption as a Function of the Amount of MetalParticles

A uniform layer of silver was deposited on a silicon dioxide substrate.The substrate was immersed in a cleaning solution of 20% sulphuric acidfor 10 minutes at room temperature, followed by rinsing in demineralisedwater. The surface of the substrate was activated by immersion in anaqueous solution of stannous chloride and the rinsed in demineralisedwater. The surface of the substrate was then plated with a uniform layerof silver by immersion in 4 baths of deposition solutions comprisingsilver ions. This yielded a silver surface with an applied amount of 0.8μg/cm² corresponding to a thickness of about 77 Å. Particles consistingof 95% palladium and 5% gold were subsequently deposited on the firstsilver surface by immersion in a dilute suspension of Pd/Au-particles.The applied amount of metal particles was 0.05, 0.12, 0.48 and 0.59μg/cm² respectively. The substrate was rinsed in demineralised water anddried.

Adsorption of fibrinogen was studied by the QCM-D technique. Fibrinogenis a glycoprotein synthesised in the liver and is found in blood plasma.QCM-D is a quartz crystal microbalance with dissipation monitoring.

The adsorbed amount of fibrinogen as a function of applied metalparticles is shown in table 2.

TABLE 2 Amount of Pd/Au-particles Fibrinogen adsorption (μg/cm²)(μg/cm²) 0.05 2.5 0.12 2.8 0.48 1.8 0.59 2.3

Example 3 Growth of Bacteria as a Function of the Amount of MetalParticles

Palladium/gold nanoparticles were deposited in different amounts on asilver base layer, following the method outlined in example 1. Theparticles comprised 95% palladium and 5% gold. The amount of silver inthe base layer was kept constant for all samples. Hence the amount ofdeposited Pd/Au particles was varied. The growth of bacteria as afunction of amount of deposited nanoparticles (Pd/Au) was studied usingthe following method:

Coated samples were placed into universals. Triplicates were includedfor each test condition 10 ml of artificial urine (AU) containinginoculated E. coli (roughly 10⁵ CFU/ml) was added to each one and theywere incubated horizontally with gentle shaking at 37° C. for 4 hours.After 4 hours the universals were removed from incubation. The sampleswere removed and CFU (colony forming unit) counts were done from eachuniversal by carrying out 10-fold dilutions in sterile distilled waterand plating 100 μl onto a third of a nutrient agar plate. These wereincubated for 16-24 hours at 37° C. and the colonies counted. Thereduction in log CFU/ml versus a control was calculated and is shown inTable 3.

TABLE 3 Amount of nanoparticles Reduction in Log CFU/ml (Pd/Au) (μg/cm²)read vs. control 0.78 6.5 0.84 7.0 1.03 6.0 1.10 6.5 1.74 5.3 2.35 4.92.41 4.6

Example 4 Microbial Growth for Several Species

Palladium/gold nanoparticles were deposited in different amounts on asilver base layer on a substrate of silicone, following the methodoutlined in example 1. The particles comprised 95% palladium and 5%gold. The amount of silver in the base layer was kept constant for allsamples. The amount of deposited Pd/Au particles was 0.36 μg/cm². Theantimicrobial properties for different bacterial strains were studied.

Species of microorganisms were chosen with the goal to survey a range ofcommon pathogens (clinical isolates) involved in bacteria transmissionand nosocomial infections, namely Escherichia coli (E. coli),Pseudomonas aeruginosa (P. aeruginosa), Enterococcus spp, Klebsiella,and Candida.

The Pd/Au coated silicone samples were placed into universals.Triplicates were included for each test condition. 10 ml of artificialurine containing inoculated organisms (roughly 10⁵ CFU/ml) was added toeach one and they were incubated horizontally with gentle shaking at 37°C. for 24 hours.

After 24 hours the universals were removed from incubation. The sampleswere removed, drained on paper towels and then placed into universalscontaining 20 ml PBS+Tween and sonicated for 1.5 minutes.

CFU counts were done from each universal by carrying out 10-folddilutions in sterile distilled water and plating 100 μl onto a third ofa nutrient agar plate. These were incubated for 16-24 hours at 37° C.and the colonies counted. In table 4 the reduction of bacteria comparedto the uncoated silicone sample is shown. The larger the value thegreater reduction.

TABLE 4 Reduction vs. Control (Log CFU/cm) E. P. coli aeruginosaEnterococcus Klebsiella Candida Uncoated 0.00 0.00 0.00 0.00 0.00silicone Pd/Au 1.64 2.53 3.88 1.37 2.52 particle coated silicone

Example 5 Primary Adhesion and Cell Recovery of P. aeruginosa

Palladium/gold nanoparticles were deposited in different amounts on asilver base layer, following the method outlined in example 1. Theamount of silver in the base layer was kept constant for all samples.The amount of Au and Pd in the particles was varied according to table5.

The samples were challenged with radiolabled P. aeruginosa GSU-3, andallowed to incubate for a period of two hours.

Primary attached cells on the samples and cell recovery was determined(ability for the cells to recover). The method employed in this studywas the one described in M. M. Gabriel et al., Current Microbiology,vol. 30 (1995), pp. 17-22, mutatis mutandis. The results are summarizedin Table 5 below.

TABLE 5 Cell Primary recovery Amount Au Amount Pd Adhesion PercentSample No (μg/cm²) (μg/cm²) CFU/mm² Reduction 1 0.08 1.17 2.45 * 10⁴ 932 0.31 0.95  2.8 * 10⁴ 95.3 3 1.01 0.56  3.2 * 10⁴ 89.9 4 1.1 0.263.45 * 10⁴ 93 5 0.98 0.02  3.4 * 10⁴ 94.4 Silicone 0 0 4.85 * 10⁴ 0Control (uncoated)

Example 6

A net of polyester fabric was first rinsed in a 5% potassium hydroxidesolution for 5 min at 30° C. After repeated rinsing in demineralisedwater the substrate was immersed in an acidified solution of 1 g/lstannous chloride at room temperature for 10 min. After rinsing indemineralised water it was soaked in a plating bath containing 2 g/lcopper sulphate, 5 g/l sodium hydroxide, 50 g/l sodium citrate and 0.005ml/l formaldehyde for 10 min at 35° C. A copper layer of about 200 Å wasobtained and after new rinsing in demineralised water the substrate wasimmersed in a particle suspension comprising 0.05 g/l each of palladiumparticles and gold particles. The applied amount of metal particles was0.4 μg/cm².

Example 7

A substrate of PMMA was cleaned in 5% hydrochloric acid for 2 min andthen rinsed in demineralised water before dipping in a solutioncontaining 0.02 g/l of the stannous ion at a pH of 2.5. After rinsingthe substrate was immersed in a solution containing 0.005 g/l of silverions, 0.02 ml/l ammonia, 0.05 g/l potassium hydroxide and 0.0005 ml/lformaldehyde for 5 min at room temperature.

This gave a surface with 0.12 μg/cm² of silver. After rinsing it wasimmersed in a particle suspension comprising 0.005 g/l palladium and0.002 g/l gold particles. The applied amount of metal particles was 0.05μg/cm².

Example 8

A non-woven polyimide substrate was immersed in a 12% solution of NaOHat 40° C. for 10 min. After repeated rinsing in demineralised water itwas immersed in an alcoholic solution containing 0.5 g/l stannouschloride for 5 min at room temperature. After rinsing it was soaked in acopper bath according to example 3. A copper layer of 2 μg/cm² wasobtained. After rinsing it was immersed in a suspension comprising 1% ofPd and 0.2% of gold particles, calculated on the weight of the totalsuspension. The applied amount of metal particles was 0.6 μg/cm².

Example 9

A nylon fabric was cleaned in 5% NaOH for 10 min at 40° C. and afterrinsing in demineralised water immersed in a solution of 0.6 g/lstannous chloride at pH 2.2 for 15 min at room temperature. After thisthe surface comprised a silver amount of 0.8 μg/cm². After a new rinsingit was dipped in a silver bath according to example 2 and then after newrinsing dipped in a suspension comprising 1% Pd and 0.05% Au particles.The applied amount of metal particles was 0.12 μg/cm².

Example 10

A substrate of aluminium was treated in a solution of 10% nitric acidand 3% hydrofluoric acid at 60° C. for 20 min. After rinsing, thesubstrate was dipped in an acidified solution of 3 g/l stannous chlorideand after renewed rinsing in a silver bath according to example 2. Afterthis step an amount of around 80 Å silver was obtained on the surface.After another rinsing the substrate was immersed in a suspensioncomprising 1% Pd and 2% Au particles. The applied amount of metalparticles was 0.7 μg/cm².

Example 11

A substrate of PTFE was etched in an aqueous solution of sodiumhydroxide for 5 min. After rinsing and drying it was immersed in asolution containing 0.7 g/l stannous chloride for 20 min at roomtemperature. The substrate was after rinsing dipped in a plating bathcontaining 0.2 g/l silver nitrate, 0.5 ml/l ammonia and sodium hydroxideto pH 10.5 for 5 min. After this step an amount of around 2.2 μg/cmsilver was obtained on the surface. After a new rinse it was immersed ina suspension comprising 3% Pd and 0.1% Au particles for 5 min at roomtemperature. The applied amount of metal particles was 0.03 μg/cm².

Example 12

A glass plate was rinsed in 10% sulphuric acid and 1% hydrofluoric acidat room temperature for 15 min. After rinsing it was immersed in a 1%stannous fluoride solution and after a new rinse immersed in a silverbath according to example 2. After this step an amount of around 140 Åsilver was obtained on the surface. After renewed rinsing it was dippedin a suspension comprising 1% ruthenium and 2% palladium particles. Theapplied amount of metal particles was 0.25 μg/cm².

Example 13

A stainless steel substrate was immersed in a solution of 15% nitricacid and 5% HF at room temperature for 30 min and then rinsed indemineralised water. The process continued following the steps inexample 11. The applied amount of metal particles was 0.9 μg/cm².

Example 14

A titanium rod was cleaned in a solution of 18% nitric acid and 2% HFfor 20 min at room temperature. The application of an electron donatingsurface and the application of metal particles was made as in example11. The applied amount of metal particles was 0.6 μg/cm².

1. A substrate comprising an electron donating surface, wherein saidelectron donating surface further comprises metal particles at an amountof about 0.001 to about 8 μg/cm² on said surface, wherein said metalparticles comprise palladium and at least one metal selected from thegroup consisting of gold, ruthenium, rhodium, osmium, iridium, andplatinum.
 2. The substrate according to claim 1, wherein said electrondonating surface is a layer of an electron donating material in anamount of about 0.05 to about 12 μg/cm².
 3. The substrate according toclaim 2, wherein said electron donating layer is a metal that is lessnoble than palladium, gold, ruthenium, rhodium, osmium, iridium, andplatinum.
 4. The substrate according to claim 2, wherein said electrondonating layer is a metal selected from the group consisting of silver,copper and zinc.
 5. The substrate according to claim 1, wherein saidsubstrate is a polymeric substrate.
 6. The substrate according to claim1, wherein said substrate is selected from the group consisting oflatex, polymers comprising vinyl groups, silicone, polyvinylchloride,polypropylene, polyurethane, polyester, copolymerisates of ethylenevinyl acetate, polystyrene, polycarbonate, polyethylene, polyacrylate,polymethacrylate, acrylonitrile butadiene styrene, polyamide, andpolyimide, or mixtures thereof.
 7. The substrate according to claim 1,wherein said substrate is selected from the group consisting of anatural polymer, a degradable polymer, an edible polymer, abiodegradable polymer, an environmental friendly polymer, and a medicalgrade polymer.
 8. The substrate according to claim 1, wherein saidsubstrate is a metal.
 9. The substrate according to claim 1, whereinsaid substrate is selected from the group consisting of stainless steel,medical grade steel, titanium, medical grade titanium, cobalt, andchromium or mixtures thereof.
 10. The substrate according to claim 1,wherein said substrate is selected from the group consisting of glass,minerals, zeolites, stone and ceramics.
 11. The substrate according toclaim 1, wherein said substrate is selected from the group consisting ofpaper, wood, woven fibres, fibres, cellulose fibres, leather, carbon,carbon fibres, graphite, polytetrafluoroethylene, andpolyparaphenyleneterephthalamide.
 12. The substrate according to claim1, wherein said substrate has the shape of a particle.
 13. The substrateaccording to claim 1 wherein the amount of the metal particles on theelectron donating surface is from about 0.01 to about 4 μg/cm².
 14. Thesubstrate according to claim 2, wherein the ratio of palladium tonon-palladium metals in said metal particles is from about 0.01:99.99 toabout 99.99:0.01.
 15. The substrate according to claim 2, wherein theratio of palladium to non-palladium metals in said metal particles isfrom about 0.5:99.5 to about 99.8:0.2.
 16. The substrate according toclaim 2, wherein the ratio of palladium to non-palladium metals in saidmetal particles is from about 2:98 to about 95:5.
 17. The substrateaccording to claim 1 wherein said metal particles, further comprise goldin addition to palladium.
 18. The substrate according to claim 1,wherein said metal particles on the electron donating surface have anaverage size of about 10 to about 10000 Å.
 19. The substrate accordingto claim 1, wherein said metal particles on the electron donatingsurface have an average size of about 100 to about 600 Å.
 20. An objectcomprising a substrate according to claim
 1. 21. The object according toclaim 20, wherein said object is a medical device.
 22. The objectaccording to claim 20, wherein said object is a disposable article. 23.The object according to claim 20, wherein said object is a dentalarticle.
 24. A method of modifying protein adsorption to an objectcomprising, applying the substrate of claim 1 to the object.
 25. Amethod of modifying bacterial adhesion to an object comprising, applyingthe substrate of claim 1 to the object.
 26. A method of reducing orpreventing bacterial growth on an object comprising, applying thesubstrate of claim 1 to the object.
 27. A method of delayingcolonisation of bacteria on an object comprising, applying the substrateof claim 1 to the object.
 28. A method of reducing or preventingtransmission of bacteria by an object comprising, applying the substrateof claim 1 to the object.
 29. A method of reducing or preventingnosocomial infections comprising, contacting a surface with thesubstrate of claim
 1. 30. A method for the manufacture of a substrateaccording to claim 1 comprising the steps: a. depositing metal particlesfrom a suspension onto said substrate, b. rinsing said substrate, and c.drying said substrate.
 31. The method according to claim 30 furthercomprising the step of depositing an electron donating material on saidsubstrate before step a.